1. Field of the Invention
The present invention relates generally to the fields of biochemistry and the stability and storage of biological samples. More specifically, the present invention relates to preservation of biological samples.
2. Description of the Related Art
Liquid biological samples may be utilized to test patients for a variety of medical conditions or measure in research subjects a variety of experimental variables in a non-invasive or minimally-invasive manner. Therefore, methods, apparatus and preservatives for collecting and storing liquid biological samples have widespread application in clinical and laboratory settings.
An example of a biological sampling method for saliva is disclosed in U.S. Pat. No. 4,774,962. A resilient absorbent inert body is chewed by a person and is subsequently introduced into a centrifuge tubule provided with an apertured floor. The centrifuged tubule is introduced into a centrifuge and subjected to a spinning process, whereupon the saliva is pressed out of the resilient body and passes through the floor into the lower part of the centrifuged tubule.
Another method and apparatus for collecting saliva from a test subject is disclosed in U.S. Patent No. 4,580,577. A flavored absorbent mass, such as a sponge, for mastication, is chewed by the subject charging the mass with saliva. The saliva is subsequently expressed from the mass. An apparatus for this method includes a barrel-piston arrangement in association with a specimen vial for storage until diagnostic testing.
Another sampling device disclosing a piston for compressing a porous mass containing saliva is U.S. Pat. No. 5,393,496. The sample container is inserted into the collection container and becomes fluidly coupled thereto by inserting the saliva collector into the sample container and pressing therein. This allows a sample of saliva to be collected, separated from the saliva collector and retained within the collection container for testing. The saliva collector includes a piston fitting closely within the sample container and a porous mass which may be compressed by the piston in the sample container to extract the sample of saliva for distribution to the collection container. A buffering solution may be retained within the sample container for mixing with the sample of saliva.
U.S. Pat. No. 5,376,337 discloses a saliva testing device including a sample container opened at one end, a tube, a piston, a holding reservoir and a filter. The piston is fixed to a first end of the tube on a first side of the piston and has a hole in fluid communication with the tube. The piston fits closely and slidably within the sample container. An absorbent pad is affixed to the piston on a second side thereof in fluid communication with the hole in the piston. The holding reservoir is in fluid communication with a second end of the tube. The filter is in fluid communication with a second end of the tube. The absorbent pad receives a sample of fluid to be tested. When the piston is slidably inserted into the sample container, a pressure is generated forcing the saliva of fluid into the holding reservoir through the tube.
U.S. Pat. No. 5,380,492 discloses a saliva sampling device including a sample container, a cap, a sample collector and a sample adequacy system. The sample container has an inner wall surface and a retaining ridge which is disposed on the inner wall surface. The sample collector includes a piece of filter paper and a holder which has a tube and a paddle coupled to the piece of filter paper. The paddle has a peripheral edge for engaging the retaining ridge of the sample container. The cap has an outer wall surface and an inner wall surface. The outer surface snugly engages the inner wall surface of the sample container. The inner wall surface has a truncated conical portion having a top and a cylindrical portion which is coupled to the truncated conical portion adjacent to the top thereof. The truncated conical portion thereof slidably engages the tube in order to guide the tube so that the cylindrical portion thereof snugly engages the tube.
U.S. Pat. No. 4,938,963 to Parnell teaches the use of citric acid and sodium benzoate to stimulate saliva production and to facilitate the use of the Yerba Santa fluid extract of the lozenge, respectively. U.S. Pat. No. 5,022,409 to Goldstein teaches an oral rinse for the collection of immunoglobulins and other substances from saliva and U.S. Pat. No. 5,103,836 to Goldstein teaches a kit and apparatus for the collection of saliva samples. However, in both of these patents of Goldstein, citric acid is used exclusively as a saliva production stimulator. Furthermore, the presence of sodium benzoate is meaningless because each device is adjusted to a final pH of 6.5 and as is well known in this art, sodium benzoate is not effective as a preservative at pH values above 4.0.
The prior art is deficient in the lack of effective means of preserving a biological sample containing analytes, e.g., proteins, lipids and carbohydrates. The present invention fulfills this longstanding need and desire in the art.
Generally, the invention features a preservative for liquid biological samples including sodium benzoate in an amount of at least about 0.15% of the sample (weight/volume) and citric acid in an amount of at least about 0.025% of the sample (weight/volume). Embodiments of this aspect of the invention may include one or more of the following features. The amount of sodium benzoate and citric acid may range between about 0.15% to about 1.00% and about 0.025% to about 0.200% of the sample (weight/volume), respectively. The preservative may be a solid. The preservative may be disposed within a vessel for storing the sample. The preservative may be coupled operatively to an absorbent mass for collecting the sample. The preservative may be coupled operatively to a substrate which may be combined with a sample after collection. The preservative may be coupled operatively to a vehicle prior to sample collection. The liquid biological sample may be saliva, tears, urine, blood, serum, plasma, sweat, vaginal fluids, semen, feces, mucous, breast milk, ascites, lymph, pleural effusion, synovial fluid, bone narrow, spino-cerebral fluid, and washings from bodily cavities (e.g., lung washings).
In general, in another aspect, the invention features a kit for preserving a liquid biological sample including a sample collecting device and a preservative coupled operatively to the device, the preservative including sodium benzoate and citric acid in an amount of about 0.15% to about 1.00% and about 0.025% to about 0.200% of the sample (weight/volume), respectively. Embodiments of the aspect of the invention featuring a kit combining a sample collecting device and a preservative coupled operatively thereto may include one or more of the following features. The sample collecting device may be a vessel. The sample collecting device may be an absorbent mass.
In general, in another aspect, the invention features a kit for preserving a liquid biological sample including a preservative carrier and a preservative coupled operatively to the carrier, the preservative including sodium benzoate and citric acid in an amount of about 0.15% to about 1.00% and about 0.025% to about 0.200% of the sample (weight/volume), respectively. Embodiments of the aspect of the invention featuring a kit combining a preservative carrier and a preservative coupled operatively thereto may include one or more of the following features. The carrier may be a substrate. The carrier may be a vehicle.
In another aspect, the invention features the combination of a vessel and a preservative disposed within the vessel, the preservative including sodium benzoate and citric acid in an amount of about 0.15% to about 1.00% and about 0.025% to about 0.200% of the sample (weight/volume), respectively. Embodiments of the aspect of the invention featuring the combination of a vessel and a preservative disposed therein may include one or more of the following features. The preservative may be a solid. The preservative may be deposited on an interior surface of the vessel. The preservative may be deposited by vacuum drying a solution including sodium benzoate and citric acid. The vessel may include an upper chamber separated by a liquid permeable partition from a lower chamber in which the preservative is disposed and further include an absorbent mass for collecting the sample, the mass being insertable into the upper chamber such that the sample may be separated into the lower chamber and mixed with the preservative. The sample may be separated by centrifugation.
In general, in another aspect, the invention features the combination of an absorbent mass for collecting the sample and a preservative coupled operatively to the mass, the preservative including sodium benzoate and citric acid in an amount of about 0.15% to about 1.00% and about 0.025% to about 0.200% of the sample (weight/volume), respectively. Embodiments of the aspect of the invention featuring the combination of an absorbent mass and a preservative coupled operatively thereto may include one or more of the following features. The mass may be impregnated with the preservative. The combination may include a vessel having an upper chamber separated by a liquid permeable partition from a lower chamber, the mass being insertable into the upper chamber such that following collection the sample may be separated into the lower chamber. The sample may be separated by centrifugation.
In general, in another aspect, the invention features the combination of a substrate and a preservative coupled operatively to the substrate, the preservative including sodium benzoate and citric acid in an amount of about 0.15% to about 1.00% and about 0.025% to about 0.200% of the sample (weight/volume), respectively. Embodiments of the aspect of the invention featuring the combination of a substrate and a preservative coupled operatively thereto may include one or more of the following features. The combination may include a vessel within which the substrate is disposed. The vessel may include an upper chamber separated by a liquid permeable partition from a lower chamber. The substrate may be disposed within the upper chamber.
In another aspect of the invention there is provided a method of preserving a liquid biological sample, in which a liquid biological sample is provided and the sample is combined with a preservative including sodium benzoate and citric acid in an amount of about 0.15% to about 1.00% and about 0.025% to about 0.200% of the sample (weight/volume), respectively. Embodiments of the aspect of the invention featuring a method of preserving a liquid biological sample may include, after the sample is provided, one or more of the following features. The method may include introducing the sample into a vessel. The preservative may be disposed within the vessel prior to introducing the sample. The preservative may be a solid. The preservative may be deposited on an interior surface of the vessel. The preservative may be deposited by vacuum drying a solution including sodium benzoate and citric acid onto the interior surface.
The method may also include, after providing the sample, collecting the sample with an absorbent mass. The preservative may be coupled operatively to the mass. The method may include, after combining the sample with the preservative, separating the sample from the mass. Separating may be accomplished by inserting the mass into a vessel including an upper chamber separated by a liquid permeable partition from a lower chamber and centrifuging the vessel so that the sample passes through the partition into the lower chamber and the mass is retained in the upper chamber. The method may include, after collecting the sample, separating the sample from the mass. Separating may be accomplished by inserting the mass into a vessel including an upper chamber separated by a liquid permeable partition from a lower chamber into which the preservative is disposed and centrifuging the vessel so that the sample passes through the partition into the lower chamber and the mass is retained in the upper chamber.
The method may also include combining the sample with a preservative which is coupled operatively to a substrate. The substrate may be disposed within a vessel. The vessel may include an upper chamber separated by a liquid permeable partition from a lower chamber. The substrate may be disposed within the upper chamber. Combining may be accomplished by introducing the sample into the upper chamber and centrifuging the vessel so that the sample passes through the substrate into the lower chamber. The method may include, after providing the sample, collecting the sample in an absorbent mass.
In another aspect, the invention features a method of preserving a saliva sample in which a preservative is introduced into the oral cavity of a mammal and a saliva sample is collected from the mammal. Embodiments of the aspect of the invention featuring a method of preserving a saliva or plasma sample may include one or more of the following features. The preservative may include sodium benzoate and citric acid in an amount of about 0.15% to about 1.00% and about 0.025% to about 0.200% of the sample (weight/volume), respectively. The preservative may be coupled operatively to a vehicle. The vehicle may be a paste, a powder, a liquid, a gel, a tablet or a capsule.
Among the following advantages of the invention may be one or more of the following. Liquid biological samples preserved according to the present invention may be stored for extended periods without refrigeration or freezing. Therefore, samples may be easily obtained and preserved for analysis in a variety of remote locations (e.g., during space travel, field research and non-clinical health care settings). In addition, vacuum drying increases significantly the shelf life of the preservative and facilitates easy storage and transportation of vessels, substrates, absorbent masses or other materials to which the preservative is coupled (e.g., by eliminating spillage problems). Moreover, because the preservative is edible it may be applied directly to an absorbent mass or introduced directly into the oral cavity of a subject from which a saliva sample is desired.